Host factors in cryptococcal pathogenesis PROJECT SUMMARY Cryptococcus neoformans is an opportunistic pathogen responsible for life-threatening disease in patients with AIDS or otherwise compromised immunity; extrapulmonary cryptococcal infection is an AIDS-defining illness. Cryptococcosis has a tremendous impact on human health, causing over one million cases of meningitis and 625,000 deaths annually worldwide, with the vast majority of deaths occurring in individuals with AIDS. The long-term goal of our studies is to define host:fungal interactions in C. neoformans pathogenesis, in order to improve both fundamental understanding of these important events and the outcome of this devastating AIDS- related opportunistic infection. This application focuses on the interactions between C. neoformans and host phagocytes, which have been implicated in fungal latency, dissemination, and virulence. Although fungal en- gulfment has been broadly described, little is known about the host factors required to mediate this process, a lack of understanding that impairs our ability to influence this interaction in favor of the host. We recently com- pleted an siRNA screen in a human macrophage-like cell line to identify kinases and phosphatases that act in C. neoformans internalization. We have now shown that two kinases, one involved in cell signaling and one in cell surface modification, are required for efficient fungal internalization b host primary cells. Mice lacking the signaling protein also show reduced dissemination of cryptococcal infection to the brain. We propose to deter- mine the mechanisms of action by which each of these proteins influences phagocytosis, and their effects on the pathogenesis of cryptococcal disease. We will use mouse models and primary cells in focused studies that exploit our expertise in biochemistry, gene expression, host biology, and post-translational modifications to de- termine mechanism. This powerful combination of approaches will elucidate crucial events of pathogenesis that determine survival and dissemination of an opportunistic microbe, and thereby the host's ability to limit disease. Our findings will also illuminate other host:microbe interactions and may suggest new directions for antifungal therapy.